During the previous project period, we developed a general procedure for evaluating the type, amount, and genetic requirements for mutagenesis by individual carcinogen-DNA adducts in mammalian cells. The salient features of the system include: (1) the chemical or biochemical synthesis of an oligodeoxynucleotide containing a well characterized carcinogen-DNA adduct (the prototype was O6-methylguanine), (ii) the insertion by using recombinant DNA techniques of the adducted oligomer into the genome of a specially constructed plasmid vector, (iii) the introduction of the adduct- containing vector into repair-proficient and -deficient mammalian cells, where the vector integrated into and replicated along with the genome of the host, and (iv) the characterization of the qualitative and quantitative features of mutagenesis induced by the adduct in the mammalian genome. In the proposed studies, we shall complete our analysis of several alkyl adducts suspected as the initiating lesions in carcinogenesis by alkylating agents and, in subsequent studies, extend the system for the analysis of the mutations induced by bulkier DNA lesions. The adducts selected for these further studies are produced by aromatic amines and vinyl halides, two classes of chemicals of known relevance in the etiology of human cancer.